Journal
BIOTECHNOLOGY AND BIOENGINEERING
Volume 114, Issue 6, Pages 1140-1150Publisher
WILEY
DOI: 10.1002/bit.26248
Keywords
matrix metalloproteinase; inhibitory antibody; deep sequencing; synthetic library; bioinformatics
Categories
Funding
- National Science Foundation [1453645]
- National Institutes of Health [R01GM115672]
- California Breast Cancer Research Program Developmental and Exploratory Award (IDEA) [21IB-0104, P200A130127]
- Div Of Chem, Bioeng, Env, & Transp Sys
- Directorate For Engineering [1453645] Funding Source: National Science Foundation
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Matrix metalloproteinase (MMP)-14 is an important target for cancer treatment due to its critical roles in tumor invasion and metastasis. Previous failures of all compound-based broad-spectrum MMP inhibitors in clinical trials suggest that selectivity is the key for a successful therapy. With inherent high specificity, monoclonal antibodies (mAbs) therefore arise as attractive inhibitors able to target the particular MMP of interest. As a routine screening method, enzyme-linked immunosorbent assays (ELISA) have been applied to panned phage libraries for the isolation of mAbs inhibiting MMP-14. However, because of suboptimal growth conditions and insufficient antibody expression associated with monoclonal ELISA, a considerable number of potentially inhibitory clones might not be identified. Taking advantage of next-generation sequencing (NGS), we monitored enrichment profiles of millions of antibody clones along three rounds of phage panning, and identified 20 Fab inhibitors of MMP-14 with inhibition IC50 values of 10-4,000nM. Among these inhibitory Fabs, 15 were not found by monoclonal phage ELISA. Particularly, Fab R2C7 exhibited an inhibition potency of 100nM with an excellent selectivity to MMP-14 over MMP-9. Inhibition kinetics and epitope mapping suggested that as a competitive inhibitor, R2C7 directly bound to the vicinity of the MMP-14 catalytic site. This study demonstrates that deep sequencing is a powerful tool to facilitate the systematic discovery of mAbs with protease inhibition functions. Biotechnol. Bioeng. 2017;114: 1140-1150. (c) 2017 Wiley Periodicals, Inc.
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